Abstract:
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O-N-acetylglucosaminylation, termed O-GlcNAcation (O-GlcNAc), is a reversible post-translational modification which presents a dynamic and highly regulated interplay with phosphorylation. New insights strongly suggest that this atypical glycosylation might be involved in striated muscle physiology, in particular in the modulation of the calcium activation properties. Thus, incubation of skinned fibers with free GlcNAc, probably via the disruption of protein-protein interaction through O-GlcNAc moieties, leads to a decrease in calcium sensitivity in skeletal as well as cardiac muscle fibers. Our aim was to investigate the effect of the increase of soleus O-GlcNAcation level via the inhibition of O-GlcNAcase, the enzyme responsible of the deglycosylation. Several regulatory contractile proteins, predominantly fast isoforms, presented a drastic increase in their O-GlcNAc level: essential and regulatory MLC, TnI and TnT. The only slow contractile protein presenting an increase of O-GlcNAc level was MLC2. Our results also demonstrated that the enhancement of O-GlcNAcation pattern was associated with an increase of calcium affinity of slow soleus fibers. Since slow fibers expressed exclusively slow contractile and regulatory proteins as assays by immunoblot analysis, this effect might involved the O-GlcNAcation of the slow isoform of MLC2. This increase of O-GlcNAcation on sMLC2 leads to the appearance of different protein pattern, corresponding to phosphorylated and/or glycosylated states of sMLC2. The potential sites of O-GlcNAcation on the slow isoform of MLC2 (Ser1 and Thr146) are different from the one identified as phosphorylated (Ser14), contributing to the hypothesis of the existence of a dynamic cross talk between phosphorylation and glycosylation which could modulate MLC2 function. Taken together, all these data strongly linked O-GlcNacation to the modulation of contractile activity of skeletal muscle.
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